Mulberry moracins: scavengers of UV stress-generated free radicals

Mulberry leaves treated with UV-C were found to accumulate three different phytoalexins, moracin C, moracin N, and chalcomoracin. The increased level of malondialdehyde in UV-treated leaves along with moracins suggested their role as a free-radical scavenger in stressed plants. All the three moracins induced under UV stress were capable of scavenging the superoxide anion generated by the xanthine-xanthine oxidase system. Also, moracins were capable of inhibiting lipid peroxidation, which strongly indicates their role as a scavenger.     

Synthesis and biological evaluation of fused oxepinocoumarins as free radicals scavengers

Some fused dihydrooxepino[f]-, [g]-, and [h]coumarins were obtained from the ring-closing metathesis of the corresponding o-allyl-allyloxycoumarins under the treatment with the first generation Grubbs’ catalyst. These compounds were tested in vitro for their antioxidant activity, and they present significant scavenging activity. They were also showed to inhibit in vitro soybean lipoxygenase.     

Synthesis and biological evaluation of fused oxepinocoumarins as free radicals scavengers

Some fused dihydrooxepino[f]-, [g]-, and [h]coumarins were obtained from the ring-closing metathesis of the corresponding o-allyl-allyloxycoumarins under the treatment with the first generation Grubbs' catalyst. These compounds were tested in vitro for their antioxidant activity, and they present significant scavenging activity. They were also showed to inhibit in vitro soybean lipoxygenase.     

Synthesis of purine homo-N-nucleosides modified with coumarins as free radicals scavengers*

Cross metathesis (CM) of 9-butenylpurines with 4-butenyloxycoumarin in the presence of Grubbs 2nd generation catalyst under MW irradiation resulted to conjugated compounds containing homo-N-nucleosides and coumarins. Analogous derivatives received by the CM reaction of 9-butenyl-6-piperidinylpurine with 6- or 7-butenyloxycoumarins, allyloxycoumarins or coumarinyl acrylate. These compounds were tested in vitro for their antioxidant activity and they present significant scavenging activity. The presence of a pentenyloxy moiety, the attachment position on coumarin ring as well as a purine homo-N-nucleoside group are considered as important structural features.     

Carotenoids as scavengers of free radicals in a Fenton reaction: antioxidants or pro-oxidants?

The spin trapping EPR technique was used to study the influence of carotenoids (beta-carotene, 8'-apo-beta-caroten-8'-al, canthaxanthin, and ethyl 8'-apo-beta-caroten-8'-oate) on the yield of free radicals in the Fenton reaction (Fe(2+) + H(2)O(2) --> Fe(3+) + .OH + -OH) in the organic solvents, DMSO, and methanol. DMPO and PBN were used as spin trapping agents. It was demonstrated that carotenoids could increase or decrease the total yield of free radicals depending on the oxidation potential of the carotenoids and the nature of the radicals. A reaction mechanism is suggested which includes the reduction of Fe(3+) to Fe(2+) by carotenoids. The effectiveness of this carotenoid-driven Fenton reaction increases with a decrease of the scavenging rates for free radicals and with decreasing oxidation potentials of carotenoids.     

Sulfur oxidation of free methionine by oxygen free radicals

The oxidation of free methionine to methionine sulfoxide by chemically or enzymatically generated oxygen free radicals is presented. The physiological significance of this process in living cells is suggested.     

Spin trapping of oxygen and carbon-centered free radicals in ischemic canine myocardium

Oxygen free radical injury has been postulated to occur during myocardial ischemia. We have used Electron Spin Resonance and Spin Trapping techniques to directly demonstrate the production of carbon-centered (R.) and oxygen-centered lipid radical (RO.) in ischemic canine heart. In addition, venous effluent from the ischemic region showed that conjugated dienes (lipid peroxidation products) increased with ischemic duration. Our results suggest that the formation of the oxygen-centered and carbon-centered lipid radical species during ischemia are a consequence of oxy-radical peroxidation of myocardial membrane lipids.     

Cell damage by oxygen free radicals

The exposure of isolated and cultured cells to oxygen free radicals generated extracellularly or intracellularly during the metabolism of foreing compounds results in the development of damage that eventually lead to cell death. Multiple mechanisms are involved in these cytopathological processes, including direct attack of free radicals to macromolecules essential for cell life, as well as indirect activation of catabolic processes such as proteases, endonucleases and phospholipases. A key role in triggering these indirect events is played by Ca²⁺ whose cytosolic concentration during oxidative stress raises well above the physiological limits.     

Influence of melatonin pretreatment and preconditioning by hypobaric hypoxia on the development of cortical photothrombotic ischemic lesion

Photothrombotic model of ischemia (PT) is based on free radical-mediated endothelial dysfunction followed by thrombosis. Free radicals are also involved in hypoxic preconditioning. We tested the sensitivity of PT to preconditioning with hypobaric hypoxia and to pretreatment with melatonin. In adult Wistar rats, after intravenous application of Rose Bengal, a stereo-tactically defined spot on the denuded skull was irradiated by a laser for 9 min. The first experimental group underwent hypobaric hypoxia three days before irradiation. In the second experimental group, melatonin was applied intraperitoneally one hour before irradiation. Three days after irradiation, animals were sacrificed, the brains perfused, and stained with TTC. Ischemic lesions were divided into grades (I, II, III). In the control group (where no manipulation preceded photothrombosis), most animals displayed deep damage involving the striatum (grade III). The group pre-exposed to hypoxia showed similar results. Only 28.57 % of the melatonin pretreated animals exhibited grade III lesions, and in 57.14 % no signs of lesions were detected. Pre-exposure to hypoxia was not protective in our model. Pretreatment with melatonin lead to a significant reduction of the number of large ischemic lesions. This result is probably caused by protection of endothelial cells by melatonin.     

The release of histamine by free radicals

An extensive discussion of the available evidence about the release of histamine by free radicals is presented. In addition, the involvement of the free-radical driven release of histamine in human pathophysiology is debated.     

Leaf senescence and lipid peroxidation: Effects of some phytohormones, and scavengers of free radicals and singlet oxygen

During in vitro senescence (chlorophyll loss) of oat ( Avena sativa L. cv. Victory) leaf segments and of leaf discs of Rumex obtusifolius L, the activity of catalase decreases and lipid peroxidation increases. The activity of superoxide dismutase (SOD) decreases in Rumex leaf discs but changes little in oat leaf segments. Kinetin treatment of oat leaf segments, and GA₃ treatment of Rumex leaf discs, inhibit decline in the enzyme activities and increase in the level of lipid peroxidation and strongly inhibit senescence. In either leaf tissue a treatment with ethanol or vitamin E (scavengers of free radicals) or with diphenylisobenzofuran (scavenger of singlet oxygen) results in a strong inhibition of lipid peroxidation and senescence, but does not affect much the decline in the SOD and catalase activities. It is concluded that, i) senscence-associated lipid peroxidation is induced by free radicals and singlet oxygen; and, ii) kinetin and GA₃ inhibit senescence mainly by a modulation of lipid peroxidation through maintaining high levels of such cellular scavengers as SOD and catalase.     

Generation of superoxide radicals by ischemic heart mitochondria

Tiron (1,2-dihydroxybenzene-3,5-disulfonic acid, disodium salt) was used as a spin trap to detect superoxide radicals produced by rat heart mitochondria. It was shown that ischemia results in the enhancement of the mitochondrial superoxide-forming activity. In the presence of the oxidative phosphorylation uncoupler mesoxalonitrile (3-chlorophenyl)-hydrazone the superoxide production rate in the control mitochondria increases, that in the ischemic mitochondria remains unchanged.     

Oxidative hemolysis of erythrocytes and its inhibition by free radical scavengers

The oxidative hemolysis of rabbit erythrocytes induced by free radicals and its inhibition by chain-breaking antioxidants have been studied. The free radicals were generated from either a water-soluble or a lipid-soluble azo compound which, upon its thermal decomposition, gave carbon radicals that reacted with oxygen immediately to give peroxyl radicals. The radicals generated in the aqueous phase from a water-soluble azo compound induced hemolysis in air, but little hemolysis was observed in the absence of oxygen. Water-soluble chain-breaking antioxidants, such as ascorbic acid, uric acid, and water-soluble chromanol, suppressed the hemolysis dose dependently. Vitamin E in the erythrocyte membranes was also effective in suppressing the hemolysis. 2,2,5,7,8-Pentamethyl-6-chromanol, a vitamin E analogue without phytyl side chain, incorporated into dimyristoylphosphatidylcholine liposomes, suppressed the above hemolysis, but alpha-tocopherol did not suppress the hemolysis. Soybean phosphatidylcholine liposomes also induced hemolysis, and a lipid-soluble azo initiator incorporated into the soybean phosphatidylcholine liposomes accelerated the hemolysis. The chain-breaking antioxidants incorporated into the liposomes were also effective in suppressing this hemolysis.     

Myocardial injuries mediated by free oxygen radicals

Dog experiments were performed to establish the lipid peroxidation of heart tissue (measured by formation of malone-dialdehyde--MDA) and the natural scavenger action (measured by determination of superoxide dismutase--SOD and of reduced glutathione--GSH). Experimental groups were: control dogs having intact heart, dogs ventilated with hypoxic gas (N2O and O2 at a ratio of 10:1) for 1, 2 and 3 hours and dogs having acute coronary ligature for 1, 2, 3 and 24 hours. Acute hypoxia caused a gradual increase of MDA concentration, a moderate increase of the GSH level and a sharp decrease in SOD activity. In ischaemic heart tissue, these changes were very distinctive. High MDA values were found after 3 hours. GSH level and SOD activity decreased continually. Increased MDA formation indicates breakdown of polyunsaturated fatty acids (PUFA) in the membranes, decreased GSH and SOD levels indicate impairment of the natural scavenging, clearly outlining the extent of disintegration of the membrane structure and function due to the effect of toxic free oxygen radicals.     

Scavenging by alginate of free radicals released by macrophages

Failure to eradicate mucoid forms of P. aeruginosa has implicated bacterial alginate in a local evasion of host defence mechanisms within the lung of Cystic Fibrosis (CF) patients. We have found that purified bacterial alginate scavenges free radicals released by triggered macrophages as detected by lucigenin amplified chemiluminescence (CL) and reduction of cytochrome c. In agreement with this, alginate was also able to scavenge radicals generated by a chemical system (hydrogen peroxide and copper; detected by benzoate hydroxylation and chemiluminescence), and by an enzymatic system (hypoxanthine and xanthine oxidase; detected by chemiluminescence). All inhibitions were dose-related. Oxygen consumption by neutrophils (unlike that of macrophages) could be detected in a Clark electrode, and was not reduced by alginate, confirming that scavenging of radicals was responsible for the earlier observations. These data suggest that bacterial alginate by scavenging free radicals, may favour the survival of mucoid forms of P. aeruginosa, particularly in the CF lung.     

Phenothiazine radicals inactivate Trypanosoma cruzi dihydrolipoamide dehydrogenase: enzyme protection by radical scavengers

Phenothiazine cation radicals (PTZ + •) irreversibly inactivated Trypanosoma cruzi dihydrolipoamide dehydrogenase (LADH). These radicals were obtained by phenothiazine (PTZ) peroxidation with myeloperoxidase (MPO) or horseradish peroxidase (HRP/H 2 O 2 ) systems. LADH inactivation depended on PTZ structure and incubation time. After 10 min incubation of LADH with the MPO-dependent systems, promazine, trimeprazine and thioridazine were the most effective; after 30 min incubation, chlorpromazine, prochlorperazine and promethazine were similarly effective. HRP-dependent systems were equally or more effective than the corresponding MPO-dependent ones. Chloro, trifluoro, propionyl and nitrile groups at position 2 of the PTZ ring significantly decreased molecular activity, specially with the MPO/H 2 O 2 systems. Comparison of inactivation values for LADH and T. cruzi trypanothione reductase demonstrated a greater sensitivity of LADH to chlorpromazine and perphenazine and a 10-fold lower sensitivity to promazine, thioridazine and trimeprazine. Alkyl-amino, alkyl-piperidinyl or alkyl-piperazinyl groups at position 10 modulated PTZ activity to a limited degree. Production of PTZ + • radicals was demonstrated by optical and ESR spectroscopy methods. PTZ + • radicals stability depended on their structure as demonstrated by promazine and thioridazine radicals. Thiol compounds such as GSH and N -acetylcysteine, l -tyrosine, l -tryptophan, the corresponding peptides, ascorbate and Trolox, prevented LADH inactivation by the MPO/H 2 O 2 /thioridazine system, in close agreement with their action as PTZ + • scavengers. NADH (not NAD + ) produced transient protection of LADH against thioridazine and promazine radicals, the protection kinetics being affected by the relatively fast rate of NADH oxidation by these radicals. The role of the observed effects of PTZ radicals for PTZ cytotoxicity is discussed.     

Effects of scavengers of oxygen free radicals on the anaerobic oxidation of 6-hydroxydopamine by H2O2

H₂O₂, a product of the aerobic autoxidation of 6-hydroxydopamine, is also consumed as a reactant, contributing progressively more to the oxidation as the concentration of O₂ becomes limiting H₂O₂ is a less effective oxidant than O₂, since the anaerobic peroxidatic oxidation of 6-hydrodopamine is slower than the aerobic oxidation by three orders of magnitude. The anaerobic peroxidation was inhibited by the hydroxyl scavengers mannitol (13–40%), glucose (41–62%) and benzoate (15–100%), implying a catalytic role for ^.OH. -9e strongly inhibitory action of desferrioxamine (76–91%), regardless of which other scavengers were present, suggests a specific role for iron in the reaction, despite the use of Chelex 100-treated buffers. Further addition of diethylenetriaminepentaacetate (DTPA), benzoate or formate to the desferrioxamine-treated reactions resulted in complete inhibition. In contrast, the presence of DTPA alone, accelerated the reaction by 160%. This acceleration is in part due to stimulation by DTPA of production of ^.OH (by Fenton-type reactions), since it was partially prevented by the hydroxyl scavengers benzoate (32% inhibition) and glucose (41%). Thus, DTPA inhibits the participation of metals other than iron, but potentiates the catalytic role of iron, in the reduction of hydrogen peroxide. The semidehydromannitol radical can reduce the DTPA-Fe³⁺ chelate directly, since mannitol further accelerated the DTPA-stimulated peroxidation (by 55%). Superoxide dismutase also accelerated the reaction (by 57–84%). This activation was seen regardless of which other scavengers were present. These effects are explained in terms of potentiating or moderating interactions among the reactive intermediates which propagate the overall reaction.